Mutations in the gene coding for Parkin cause a rare familial form of Parkinsonism, autosomal recessive juvenile Parkinsonism, that results in death of dopaminergic neurons in the substantia nigra. To understand how parkin causes disease, we need to understand the regulation and function of parkin. Our studies have lead us to investigate ubiquitination, which is a process that regulates protein degradation. We hypothesize that parkin regulates ubiquitination of other proteins in response to cellular contact with matrix proteins (such as collagen and laminin), and thereby controls regulation of the cytoskeleton and signal transduction by matrix proteins and their integrin receptors. Loss of parkin function could cause neurodegeneration by inhibiting matrix signaling and impairing maintenance of processes by neurons. Our preliminary data support this hypothesis by demonstrating that parkin-dependent ubiquitination is activated by cellular binding to matrix proteins. We have also identified parkin binding proteins that are associated with integrins. Conversely, cell lines that have reduced parkin expression (due to anti-sense parkin cDNA) decrease ubiquitination, retract processes upon cellular exposure to matrix proteins, and have abnormal signal transduction. The goal of this proposal is to investigate the regulation of ubiquitination by parkin (Aim 1), determine the role of parkin in regulating signaling in response to exposure of cells to matrix proteins (Aim 2) and identify common functional deficits associated with disease-related mutations in parkin. Interestingly, parkin is also linked to other forms of neurodegeneration. Parkin binds to alpha-synuclein, and in brains from donors with Parkinson's disease parkin accumulates in inclusions that contain alpha-synuclein, and shows 75% less binding of parkin to two proteins, filamin and hCDCrel2a. We intend to investigate the mechanism of parkin dysfunction by determining how parkin function is altered in Lewy body diseases, and whether oxidation or alpha-synuclein aggregation causes the dysfunction of parkin (Aim 3). The research in this proposal will provide insight into the function of parkin, determine how mutations in parkin produce disease, and provide a new window to understand the molecular pathophysiology of Parkinson's disease.